Hepatitis C Virus Protease Inhibitors Show Differential Efficacy and Interactions with Remdesivir for Treatment of SARS-CoV-2 In Vitro

Abstract

Antivirals targeting SARS-CoV-2 could improve treatment of COVID-19. We evaluated efficacy of clinically relevant hepatitis C virus (HCV) NS3 protease inhibitors (PI) against SARS-CoV-2 and their interactions with remdesivir, the only direct-acting antiviral approved for COVID-19 treatment. HCV PI showed differential potency in short-term treatment assays based on detection of SARS-CoV-2 Spike protein in VeroE6 cells. Linear PI boceprevir, telaprevir and narlaprevir had 50% effective concentrations (EC50) of ∼40 μM. Among macrocyclic PI, simeprevir had the highest (EC50 15 μM) and glecaprevir the lowest (EC50 >178 μM) potency, with paritaprevir, grazoprevir, voxilaprevir, vaniprevir, danoprevir and deldeprevir in between. Acyclic PI asunaprevir and faldaprevir had EC50 of 72 and 23 μM, respectively. ACH-806, inhibiting the HCV NS4A protease cofactor, had EC50 of 46 μM. Similar and slightly increased PI potencies were found in human hepatoma Huh7.5 cells and human lung carcinoma A549-hACE2 cells, respectively. Selectivity indexes based on antiviral and cell viability assays were highest for linear PI. In short-term treatments, combination of macrocyclic but not linear PI with remdesivir showed synergism in VeroE6 and A549-hACE2 cells. Longer-term treatment of infected VeroE6 and A549-hACE2 cells with 1-fold EC50 PI revealed minor differences in barrier to SARS-CoV-2 escape. Viral suppression was achieved with 3- to 8-fold EC50 boceprevir or 1-fold EC50 simeprevir or grazoprevir, but not boceprevir, in combination with 0.4- to 0.8-fold EC50 remdesivir; these concentrations did not lead to viral suppression in single treatments. This study could inform development and application of protease inhibitors for optimized antiviral treatments of COVID-19.

In vitro efficacy of artemisinin-based treatments against SARS-CoV-2

“…In these experiments the Danish SARS-CoV-2 isolate SARS-CoV-2/human/Denmark/DK-AHH1/2020 was used employing a 96-well plate based concentration-response antiviral treatment assay, allowing for multiple replicates per concentration, as described in "Materials and methods" and Supplementary Information (Figures S3 and S4) 20,21 . Seven replicates were measured at each concentration and a range of concentrations was evaluated to increase data accuracy when compared to the plaque-reduction assay, which was carried out in duplicates.…”

Efficacy of Ion-Channel Inhibitors Amantadine, Memantine and Rimantadine for the Treatment of SARS-CoV-2 In Vitro

“…African green monkey kidney VeroE6 cells, human hepatoma Huh7.5 cells and human lung carcinoma A549-hACE2 cells were cultured as described [ 13 , 14 , 15 ].…”

“…SARS-CoV-2 infected VeroE6 cultures were treated immediately after virus inoculation and upon cell splitting [ 13 , 14 , 15 ]. Cultures were evaluated by immunostaining for SARS-CoV-2 spike protein upon cell splitting.…”

“…VeroE6, Huh7.5 and A549-hACE2 cells in 96-well plates (Thermo Fischer Scientific, Roskilde, Denmark) were inoculated with SARS-CoV-2 at MOI 0.002, 0.02 or 0.003, respectively, and treated with inhibitor dilutions. MOI for these assays were selected based on virus titration assays carried out in the different cell lines as described [ 14 ], to obtain fully infected cultures whilst avoiding virus-induced cytopathic effects. Treatment conditions were tested in 7 replicates.…”

“…Single SARS-CoV-2 spike protein positive cells were automatically counted using an ImmunoSpot series 5 UV Analyzer (CTL Europe GmbH, Bonn, Germany). Counts from infected-treated wells were related to the mean count of infected-nontreated wells to calculate the %residual infectivity for single inhibitor treatments and, in addition, the % inhibition for combination treatments and time-of-addition experiment [ 13 , 14 , 15 ].…”